Skin, the outermost layer of mammals, acts as a barrier to protect the body from hazards in the environment and to keep fluids within the body. Skin originates from a single layer of embryonic skin progenitor cells that give rise to skin lineages: the epidermis and hair follicles. In an adult organism, separate pools of epidermal and hair follicle stem cells control skin maintenance throughout the lifespan of the animal. Identification of the molecular mechanisms that control skin morphogenesis during embryonic development, as well as those that control the self-renewal and differentiation of skin stem cells during adulthood will enhance our knowledge of skin biology and of the basis behind various skin disorders. My previous work revealed that a key epigenetic regulator, the Polycomb complex, represses the epidermal differentiation program and promotes the proliferation of embryonic skin progenitor cells. The central core of this proposal is to elucidate the roles of the Polycomb complex in the control of embryonic and adult skin stem cells. During the K99 phase, in vivo loss-of-function studies will uncover whether the Polycomb complex is important for the establishment and maintenance of skin lineages by skin stem cells. Next, in order to characterize the mechanism of action of this complex, biochemical approaches will be used to identify Polycomb target genes. During the R00 phase, the roles of identified targets in the control of stem cell maintenance and differentiation will be examined. Completion of these studies will reveal the role of the Polycomb complex in skin control and will also establish a paradigm for future studies of other epigenetic regulators. Recently, my studies have revealed differential expression patterns of additional important epigenetic regulators in embryonic skin cell progenitors and differentiated cells, suggesting an important role for epigenetic regulation in the control of skin stem cells. To characterize this epigenetic regulatory network, the roles of these epigenetic factors in the control of epidermal development will be examined during the R00 phase. In vitro functional studies will reveal whether the selected epigenetic regulators control proliferation and differentiation of epidermal cells. The in vitro findings will be next examined in vivo by looking for epidermal reconstitution upon in utero injections of viruses that alter expression of the candidate genes or by engraftment of overexpressing and/or knock-down cells. Finally, rigorous biochemical and molecular studies will uncover the mechanism of action of the epigenetic factors. In summary, the completion of these studies will unveil the roles of epigenetic regulators in the control of a tissue-skin-that is essential for our survival.